Use of Combination Preparations Comprising Antifungal Agents

ABSTRACT

The invention relates to the use of an antimycotic agent and an epithelial cell or endothelial cell adhesion inhibitor for producing a combination drug for the topical treatment of  Candida  mycoses selected from vulvovaginal candidiasis, oropharyngeal candidiasis (oral thrush), diaper dermatitis (diaper thrush) and intertriginous ( Candida ) eczema.

The invention relates to the use of combination preparations, comprisingantimycotics, for the treatment of Candida mycoses. Yeast, primarilyCandida spp. (Candida albicans) usually are opportunistic pathogenswhich become pathogenic only when the systemic or the local immunedefence is impaired (e.g. oral thrush in HIV patients, leukemiapatients, under chemotherapy, in babies; intertriginous eczema inDiabetes mellitus). An exception is vulvovaginal Candida mycosis of anotherwise healthy woman. However, also with the vaginal candidose thereare a number of factors which cause an increase in the virulence of thepathogen, and which, in case of a respective personal disposition, maychange from an asymptomatic colonization to a symptomatic disease.

The vulvovaginal mycosis is a very frequent disease which hascontinuously increased over the last years. According to recentestimates, three out of four women get a vaginal mycosis at least oncein their lifetime. In 5% of the cases, the disease takes achronic-recurrent course which often constitutes a severe subjectiveimpairment for the afflicted patient. Treatment of the vaginal mycosisis particularly difficult during pregnancy, since in this case thereexist particularly promoting factors on the side of the patient, whileon the other hand the possible therapies are considerably restrictedand, thirdly, a therapy is absolutely necessary since otherwise thenewborn baby will be infected during birth.

Oro-phyryngeal thrush diseases in turn pose a problem not only innewborns with their immature immune system, but also in patients with animpaired immune defence, such as HIV patients, patients undergoingchemotherapy and patients suffering from malignant hematologicaltumours. As has been mentioned, it is known that infectious diseasesattack debilitated and immunosuppressed humans particularly easily,wherein—as is the case in many diseases caused by Candida ssp.—harmlesssymbionts become pathogens.

Nevertheless, this fact has not been sufficiently mechanisticallyconsidered in the therapeutic concept. So far, no drug or combination ofpharmaceutical substances in a drug has received a market authorizationby means of which the infectiousness of the pathogen on cell level hasdeclaredly been reduced or stopped by influencing physiologicalreactions of the patient in combination with a direct “attack” on thefungus.

At present, the therapy of cutaneous and mucocutaneous candidiasis isexclusively effected by antimycotics which, depending on the respectivedose, have a fungistatical or funcidal effect and, depending on theactive substance, are applied systemically or locally.

For the therapy of candida mycoses, such as the thrush of the oralcavity or the vaginal mycosis, a plurality of drugs having anantimicotic effect has been granted market authorization, in most casesfor the topical, yet also for the systemic therapy. The mechanism of thepharmacologic activity of nearly all the pharmaceutical substances usedis well known. Obviously, most of the known mechanisms are geared to theinfective organism (bacterium or fungus) itself. Thus, a number of thesepharmaceutical substances interferes in the ergosterole-biosynthesis(isoprene biosynthesis pathway) by inhibition. The pharmaceuticalsubstance attempts to inhibit or kill the fungus via physiologicalprocesses that should differ from the human metabolism as far aspossible.

In WO 00/56353 A2, peptides derived from alpha-metanocyte-stimulatinghormone have been suggested for the treatment of Candida vaginitis.

EP 1 637 132 A1 relates to compositions for the treatment of athlete'sfoot in sportsmen.

EP 0 592 348 A1 discloses compositions for the treatment of ophthalmicdiseases.

WO 2005/117831 describes posaconazole-containing preparations forinjection for the treatment of mycoses.

According to WO 00/48633 A1, antibodies to hydrophobic proteins havebeen suggested for combating Candida mycoses.

US 2003/181384 A1 relates to administering trefoil peptides forpreventing epithelial damage possibly caused by infections. Localanti-inflammatory vaginal rinses containing benzydamine for themechanical removal of a mass of fungus have been suggested by Levy (Rev.fr. Gynecol. Obstet. 84 (1989), 779-781).

For Candida vaginitis, primarily in case of a chronic course thereof,the available spectrum of active agents does not yield any satisfactoryoption of therapy. This is particularly true if the disease occursduring pregnancy.

The molecular factors underlying the personal disposition are not knownin detail, and accordingly, so far there is no drug, or combination ofpharmaceutical substances in a drug, with which the infectiousness ofthe pathogen on cell level is reduced, or stopped, respectively, by aconcomitant direct “attack” on the fungus and an influence on thephysiological reactions of the patient. Therefore, there is an urgentdemand for therapies against Candida mycoses which so far cannot besuccessfully treated by administering conventional antimycotics. Inparticular, Candida mycoses shall be treated which relate to infectionsof the mucosa, on the one hand, mainly vulvovaginal candidiasis,oropharyngeal candidiasis (oral thrush), and, on the other hand, Candidamycosis of previously damaged (macerated) skin, such as diaperdermatitis (diaper thrush), and intertriginous eczema. Such a therapyshould also enable treatments of vulvovaginal mycoses in pregnant womenwhich requires a particularly careful use of drugs.

Accordingly, the present invention relates to the use of

-   -   an antimycotic agent and    -   an inhibitor of the adhesion of epithelial cells or endothelial        cells, for producing a combination drug for the topical        treatment of Candida mycoses selected from vulvovaginal        candidiasis, oropharyngeal candidiasis (oral thrush), diaper        dermatitis (diaper thrush) and intertriginous eczema.

The mycoses to be preferably treated according to the invention have incommon that not only the pathogen, but also the afflicted organism (ofthe patient) contributes considerably to the generation of the diseaseand to the maintenance of the infection. Apart from the fact that thetendency to contract a disease is basically higher in patients having animpaired immune system, it is the mechanisms of infectiousness which areof particular relevance in this connection. Only in recent years, suchconcepts have increasingly been investigated in the research regardinganti-infective agents.

The first step determining the infectiousness when a cell is afflictedby a pathogen consists in the pathogen's attachment to the plasmamembrane of a cell. In general, a plurality of different systems ofadhesion molecules, i.e. cell-cell-interactions, have become known forsuch an attachment. The general ability of pathogens for adhering toepitheliums and edotheliums also occurs via the interaction withadhesion molecules. For certain pathogens, for certain organs and forcertain clinical pictures of diseases, a special set of involvedadhesion molecules is to be expected. For infections in general, andmycotic infections in particular, however, adhesion factors which areonly partially involved have been published, without, however, havingbeen summarized in a comprehensive concept, or without having beenintroduced into a therapeutic concept. The present invention is based onthe finding that the same adhesion molecules which play a decisive partin the course of the coagulation of blood during the adhesion of thethrombocytes and whose expression is induced by derivatives of thearachidonic acid metabolism (inter alia von Willebrand-factor,vitronectin, fibronectin, integrins), are induced during thetransformation to pathogenicity by accompanying factors or the pathogenitself and, subsequently, are utilized for the adhesion by the primarilyopportunistic pathogens.

It has been known for quite some time that under the influence ofprostaglandins, Candida fungi undergo a transformation of their shapefrom budding shape to hyphae shape which is accompanied by an increasedgrowth. Prostaglandins are derivatives of arachidonic acid. The activeprinciple of the class of pharmaceutical substances of the NSAIDs, whichare mainly used for the treatment of inflammatory and rheumaticdiseases, primarily consists in the inhibition of the prostaglandinbiosynthesis. The influence of NSAIDs on the growth of fungi has beendemonstrated in in vitro experiments (Scott et al., Antimicrob. Ag.Chemother. 39 (12) (1995), 2610-2614; Tariq et al., Antimicrob. Ag.Chemother. 39 (12) (1995), 2615-2619; Yucesoy et al., J. Chemother. 12(2000), 385-389). In this combination with anti-mycotic agents, anincrease in the anti-mycotic effect has occasionally been observed, yetthe term “paradox effect” has been explicitly established for this,since, in vitro, at some concentrations and in some combinations,instead of an inhibition there results an increased growth (Arai et al.,Mycoses 48 (2005), 38-41). Thus, the effects of Ibuprofen in combinationwith various antimycotics have been very varied (Tariq et al., 1995).Fluconazole-sensitive strains of C. albicans have not shown an improvedresponse to a combination of Ibuprofen with Fluconazole in the invitro-system (Arai et al., 2005). In summary, the results of thein-vitro tests have shown that, based on the growth of the fungus, theeffects depend on the respective fungus strain used, yet that from theseno conclusions can be drawn on the medical efficacy at the site of themycotic infection, neither in the one (positive) nor in the other(negative) direction.

Since usual in-vitro experiments regarding an anti-mycotic activity ofcertain substances or substance combinations do not take thephysiological response of the host into consideration, in none of thesein-vitro projects, the part which the cell-cell adhesion plays in theinfection in general, or changes occurring in the host's epithelium orendothelium, respectively, can have been addressed in the least.

Moreover, in the relevant literature, the therapy resistance occurringin the treatment of chronic-recurrent Candida vulvovaginitis i.a. hasbeen attributed to the increased occurrence of certain Candidasubspecies (without, however, offering a conclusive total concept). Yet,especially for these subspecies, no in vitro-results whatsoever areavailable.

In case of Candida, in fact the transformation of the growth habit fromthe budding form to the hyphae form is a prerequisite for the layer-typeovergrowth. This clearly intensifies the pathogenicity of the pathogen.The formation of the hyphae in turn is promoted by prostaglandin E2which is also formed by the patient's endothelial cell/epithelial cellduring the inflammatory process. Also the increased mucus formation ofmucous membranes occurring during the inflammatory process plays a partin this pathological process.

Under certain promotive conditions in the microenvironment, such as,e.g., an impaired immune situation, yet also in case of suitablehormonal conditions or a patient's genetic disposition or, on the otherhand, due to quorum sensing, some pathogens, such as, in particular alsoCandida species, are capable of overgrowing the affected sector of theskin, mucosa or endothelium by forming a closed layer. This not onlyaggravates the pathological process; due to the transformation of thegrowth habit, also the pathogen's vulnerability by local treatmentmethods changes. In this growth habit, the pathogens practically can nolonger be attacked by the usual locally applied anti-mycotic agents.Chronification occurs, and it becomes necessary to change from a localtherapy to a systemtic therapy. A systemic antimycotic therapy, however,often constitutes a great strain due to the spectrum of side effects ofmany antimycotic agents and, in case of pregnant women, it is completelyimpossible because of teratogenic effects. Yet, even under systemictherapy, the chronic-recurrent vulvovaginitis and colpitis caused byCandida spp. (with Candida albicans and Candida glabrata being the mostfrequent pathogens) often are not cured, and many female patients sufferfrom several recurrences each year.

The present invention is based on the so far unknown finding which,however, is central with regard to infectiousness that, when attachingto epithelial cells (or to endothelial cells, respectively), yeast fungiuse arachidonic-acid-based mechanisms of the body itself, by which theorganism usually reacts to cell-damaging noxae.

Arachodonic acid is known to be converted on the cyclooxygenase pathwaynot only to prostaglandins, but also to thromboxane and prostacycline.Under the influence of thromboxane, the thrombocytes aggregate andadhere to the damaged membrane so as to seal the latter. In order toenable an adhesion, different adhesion molecules are not only presentedby the thrombocyte; also the endothelium specifically prepares itselffor the adhesion. On molecular level, these effects are primarilymediated via adhesion molecules, such as, e.g., von Willebrand factor.The processes taking place during the adhesion occur temporally adjustedin the thrombocyte and in the vessel endothelium. It is mainly twoderivatives of the arachidonic acid cascade which play a decisive roleas adversaries in the process of blood coaguation: besides thromboxanewhich causes the aggregation of thrombocytes, it is the “adversary”prostacyclin which prevents the reactive process from spreadingexcessively. The biosynthesis of thromboxane and prostacyclin ismediated by different enzymes.

Thromboxane is synthesized via cyclooxygenase-1 (COX1), prostacyclin viacyclooxygenase-2 (COX2).

An essential element of the mycotic infection consists in that thefungus/host interaction comprises a series of mechanistic analogies tothe thrombocyte/endothelium interaction. Primarily, this means thatinflammatory processes occur in the host cells either before orsimultaneously with the mycotic infection, i.e. the transformation ofapathogenic, or possibly pathogenic, respectively, to pathogeniccausative agent (e.g. Filler et al., Inf. Immun. 64 (1996), 2609-2617;Cannom et al., J. Inf. Dis. 186 (2002), 389-396), which are controlledvia the arachidonic acid cascade, or that the adhesion of the fungus isrendered possible at all by previous inflammatory processes in the hostcell. On the part of the fungus, adhesion of the fungi to the host cellsoccurs via the hyphae, on the part of the host cell, it is substantiallyeffected via an arachidonic-aciddependent mechanism, utilizing theadhesion molecules expressed under these conditions at the host cell,such as, e.g., von Willebrand factor, vitronectin, fibronectin orvarious integrins.

Providing an epithelial cell or endothelial cell adhesion inhibitor inthe combination preparation according to the present invention is,therefore, an essential element for the anti-mycotic agent to becomeeffective at all in the infected area of the Candida mycoses to betreated according to the invention. The nature of the epithelial cell orendothelial cell adhesion inhibitor as such is not critical, theselection is generally dictated by galenic aspects for (promoting an)optimal activity of the antimycotic agent. Therefore, antimycotic agentand epithelial cell or endothelial cell adhesion inhibitor are optimizedin the combination preparation also on the basis of theirphysico-chemical properties. This adhesion inhibitor according to theinvention exerts an influence on the arachidonic acid metabolism so thatthe inventive combination is capable of synergistically acting on themycotic infection at the site of infection and by utilizing thepatient's endogenous arachidonic acid mechanisms.

Preferably, therefore, the epithelial cell or endothelial cell adhesioninhibitor is selected from

-   -   a non-steroidal antiphlogistic agent (NSAID) with a sufficient        COX1 inhibition which, on the one hand, results from the        intrinsic activity and, on the other hand, from the extent of        the selectivity of the inhibition of COX1 relative to COX2 under        therapeutic conditions. This means that all the substances which        exhibit a more selective COX2 inhibition than, e.g., Meloxicam        or Diclofenac, are not suitable as defined by the invention        (such as, of course, also all the substances which do not        exhibit any activity or effects on the arachidonic acid        metabolism). The selection in terms of number is effected, e.g.,        by stating the COX1/COX2 ratio of the IC50 values, yet this will        differ with the method used. Suitable compounds may, e.g., be        defined in terms of numbers in accordance with the data supplied        by K. Brune et al. (Deutsches Ärzteblatt 97, (26) 2000,        A-1818/B-1538/C-1434) by a COX1/COX2 ratio (IC50) of ≦20,    -   prostacyclin or a prostacyclin analogue, preferably Ilaprost or        Cicaprost; or    -   inhibitors of the expression of epithelial- and endothelial        adhesion molecules, preferably Ticlopidin or Clopidogrel.

These three categories constitute the main inhibitors of epithelial cellor endothelial cell adhesion. For all the three groups, substances atpresent already have a market authorization or are in an advanced-stageclinical study.

Therefore, in the combination preparation according to the inventiondrugs are preferably contained which inhibit the thrombocyteaggregation, such as mainly NSAID with COX1-inhibition, or medicamentswhich prevent the expression or the function of adhesion molecules inthe endothelium connected with this process, which exacerbate or preventthe adhesion of pathogens to the endothelium or epithelium, respectivelyin an analogous manner. This is not only effected via the suppression ofthe expression of suitable adhesion molecules, but mainly also bytriggering/increasing the shedding of these surface molecules. In thesame way, prostacyclin itself and prostacyclinderivatives are capable ofreducing or stopping the adhesion. The inventive use of such compoundsin combination with an anti-mycotic agent thus both prevents theinfection, and also prevents the “boosting” of the infection triggeredby inflammatory processes occurring in the course of the infection. Thefinding that such a complex process as the adhesion of a fungus to theendothelium of the host can be influenced by active substances of awell-known class has not been known so far and forms the basis of thecombination preparations according to the invention by which thesurprising clinical results according to the present invention could beachieved, in particular in case of the preferred Candida mycosesdescribed which so far could not be treated or could be treated onlywith difficulty.

The inventive combinations of pharmaceutical substances are intended fortopical application (at mucosae or also on highly inflamed skin areas,such as in case of diaper thrush, and intertriginous eczema) fortreating mycoses (vulvovaginitis oropharyngeal candidiasis (oralthrush), diaper dermatitis intertrignious eczema) and, as mentionedbefore, consist of an anti-mycotic agent and an adhesion inhibitor ofepithelial cells or endothelial cells, which inhibitor exacerbates, orprevents, respectively, the adhesion of the pathogen to the epithelialand/or endothelial cells and the formation of a layer-shapedcolonization of the affected epithelial or endothelial sector, bysuppressing the above-described mechanisms.

As has been mentioned above, it is particularly preferred to use anNSAID compound. Yet, not all the NSAIDs are suitable compounds in thecombination preparation according to the present invention, sinceNSAIDs, depending on their profiles (COX1-inhibitor or COX2-inhibitor),generally have a promoting (COX2) or inhibiting effect (COX1) on anadhesion. For this reason, the treatment of dermatologic diseases with acombination that contains a selective COX2-inhibitor (such as, e.g.,disclosed in US 2005/0014729 A) is wrong with regard to the therapeuticaim of the present invention, i.e. causing the reduction of the adhesionin the patient's endothelium or epithelium. Highly selective COX2inhibitors or anti-inflammatory agents without an activity on thearachidonic acid mechanism (such as, e.g., benzydamine (Riboldi et al.,Br. J. Pharmacol. 140 (2003), 377-383)) are no relevant compounds asdefined by the present invention.

The selectivity of the NSAID is, e.g., expressed by the ratio of the IC50 (microM) COX1/COX2, yet the values will generally depend on thepharmacological test systems used and will differ greatly, andtherefore, according to the invention, the values are determined interms of numbers according to the data given by K. Brune et al.(Deutsches Ärzteblatt 97, (26) 2000, A-1818/B-1538/C-1434).

What is essential for the therapeutic effectiveness is a sufficientCOX1-inhibition (absolute value of the IC 50) at the respectiveadministered therapeutic dose. Most of the NSAIDs are mixedCOX1/COX2-inhibitors. As long as there exists a COX1 inhibition, theCOX2-inhibition is very welcome (because of the suppression of pain).Therefore, NSAIDs without or with only a slight preference of the COX2are substances to be used according to the invention. Not suitable are,however—as mentioned before—(highly) selective COX2-inhibitors which donot have a COX1 effect or have only an insufficient COX1 effect undertherapeutical conditions (dosages). On the contrary, the selective COX2inhibitors may even negatively affect the clinical picture and,therefore, are to be avoided within the scope of the therapy accordingto the invention. Therefore, according to the invention NSAIDs whichhave a value of >20 for the COX1/COX2-ratio (based on the above-citedliterature) are to be excluded (for highly-selective COX2-inhibitorswith a market authorization, values of more than 100 result, e.g.Celecoxib, Rofecoxib, Valdecoxib, Etoricoxib).

Accordingly, the NSAID in the combination preparation according to thepresent invention preferably is selected from Diclofenac, Fenclofenac,Alclofenac, Lonazolac, Clidanac, Oxipinac, Clopinac, Tolmetin,Indomethacin, mefenamic acid, flufenamic acid, meclofenamic acid,tolfenamic acid, niflumic acid, Floctafenin, bucloxic acid, Ibuprofen,Dextroprofen, Prapoprofen, Miroprofen, Fenoprofen, Fluprofen,Flurbiprofen, Ketoprofen, Alminoprofen, Tioxaprofen, tiaprofenic acid,Isoxepac, Nimesulid, Meloxicam, Tenoxicam, Lornoxicam, Piroxicam,Droxicam, Sudoxicam, Naproxen, Bufexamac, Etofenamate, Felbinac,Nabumeton, Ketorolac, Etodolac, Oxaprocin, salicylic acid,acetylsalicylic acid, Flufenisal, Diflunisal, Benorylat, Fentiazac,Azapropazon, Phenylbutazone, Kebuzon, pharmaceutically active salts oresters of these substances or mixtures of these substances, inparticular Diclofenac, Nimesulid, Fenclofenac, Alclofenac, Lonazolac,Tolmetin, Indometacin, mefenamic acid, flufenamic acid, meclofenamicacid, Floctafenin, Ibuprofen, Flurbiprofen, Ketoprofen, Alminoprofen,Meloxicam, Tenoxicam, Lornoxicam, Naproxen, Etofenamate, Felbinac,Nabumeton, Ketorolac or Etodolac.

The particular significance of the combination according to theinvention of an antimycotic agent and an adhesion inhibitor ofepithelial or endothelial cells will directly depend on the extent ofthe effect of the second component which primarily contributes to thetherapeutic effect by an adhesion-suppressing effect on the patient'sendothelium and/or epithelium. Therefore, it is of particular importanceto consider that the term “skin diseases” covers a broad range of skintissues of completely different structures: from an adult's nail to ababy's mucosa. What is common to all the diseases which have beenmentioned as preferred indications for the combination preparationaccording to the present invention (vulvovaginal candidiasis,oropharyngeal candidiasis (oral thrush), diaper dermatitis (diaperthrush) and intertriginous eczema) is that it is the surface of theafflicted tissue sectors which, due to the expression of correspondingadhesion molecules, enables an adhesion of the fungal colonies. Thecombination preparation according to the invention serves for thetherapy of mycoses, preferably for the therapy of Candida mycoses which,typically, occur on the aforementioned skin types, i.e. mucosae of thegenitourinary tract and the oropharyngeal region as well as onpreviously damaged (macerated, injured, inflammatorily changed) outerskin.

Accordingly, it is not a subject matter of the present invention tocarry out a therapy of dermatophyte diseases (EP-1 390 031-B1) with thecombination preparation described, which diseases often colonize thekeratic layers of the epidermis, since there the contribution to thetherapeutic success which is provided by preventing the adhesion of thefungus to the host's epithelium is markedly lower and, therefore, theinventive effect will not occur. This also holds, e.g., for thetreatment of mycoses of the nails (WO 2000/028821 A1), since here, too,the use of the inventive combination preparation is not purposefulsince, here, too, the surprising synergism of the two components in theinventive preparation will not occur (to the inventive extent), inparticular because in this instance the state of the epithelium is ofminor importance compared to the particularities of the therapy of thesediseases, in particular the poor “accessability” of the therapeuticagent.

For the topical therapy according to the invention with the describedcombination drug on mucosae or previously damaged outer skin,respectively, according to the invention not only significantly lowerconcentrations of the active substance will basically suffice ascompared to diseases of the normal or more keratinized outer skin andits adnexa, also a significant improvement of the subjective andobjective disease symptoms will occur within a substantially shortertime (from minutes to a few hours vs. from days to weeks when applyingthe treatment methods common at present). Thus, the particularly lowdosage of the adhesion-inhibiting active substance is a substantialadvantage with the present invention. This rapid start of theeffectiveness is solely caused by the procedures on the skin or mucosalsurface that has been changed due to inflammation and, therefore, can beforecast or determined in principle neither by in vitro-tests nor byexaminations on the non-inflamed outer skin. Since the greatly enhancedeffect is caused solely based on mechanisms occurring on the exposedsurface of the respectively afflicted organ (expression and rejection ofthe adhesion sites), particularly also this effect cannot be determinedby in-vitro experiments, and, in fact, it has been shown that in-vitroexperiments, depending on the fungus strain used, the dosage and thecombinations of pharmaceutical substances used, have yielded the mostcontradictory results. In this context, it is also notable that thecombination of Fluconazol with Iboprofen in Fluconazol-sensitive strainshas not shown any synergistic effect in the in-vitro experiment (Arai etal., 2005). The effect obtainable with the combination preparation ofthe invention therefore also cannot be derived in this way (via in-vitroexperiments). Also in case of a systemic administration of the inventivecombination preparation, in particular of the epithelial cell orendothelial cell adhesion inhibitor (i.e., e.g., the NSAID chosenaccording to the invention), for pharmacokinetic reasons a respectiveeffect is not to be expected even at a common dosage, and much less soat the low dosages which are made possible with the present invention.

The required low dose is also due to the fact that the epithelial cellor endothelial cell adhesion inhibitor exerts its effect both on thefungus and on the epithelium of the patient.

Thus, also a treatment of breastfeeding babies and of pregnant women ispossible.

Thus, the effect of the inventive combination of pharmaceuticalsubstances is based on

1. suppressing pathogen growth,2. suppressing adhesion of the pathogen on the host cell,3. suppressing the acute inflammation and pain symptoms by inhibitingthe prostaglandin synthesis on the part of the afflicted organism,4. preventing the generation of the promotive factors in the localenvironment, i.e. the adhesion to the host cell upon transformation ofthe pathogen and, thus, preventing an aggravation of the clinicalpicture, and5. interrupting the pathogenetic mechanism in case anaggravation/chronification has occurred,wherein 2.-5. are mediated by the epithelial cell or endothelial celladhesion inhibitor (which interferes in the arachidonic acid cascade)and 1. is mediated according to the mechanism of activity of theantimycotic agent used.

This allows for a therapy concept which is completely new as compared tothe prior art, which—apart from the effectiveness which by itselfalready is surprising in (chronic-recurrent) vulvovaginal candidiasis,oropharyngeal candidiasis (oral thrush), diaper dermatitis (diaperthrush) and intertriginous eczema—is, moreover, characterized by thefollowing advantages and effects compared to the prior art:

1. possibility of a local therapy,2. in case of chronification, marked reduction of the tendency torecurrence,3. highly accelerated onset of effect,4. significantly reduced dosage of the antimycotic agent, and5. immediate pain relief.

Ad 1. In case of promotive factors, in particular also for anaggravation/chronification, e.g. an immunosuppression, hormonal andgenetic factors, according to the present invention the pathogeneticmechanism on which the disease disposition is based is inhibited. Thedisease can be controlled, and cured, respectively, without a systemictherapy. Avoiding the changeover to a systemic therapy (which often iseffected by using pharmaceutical substances that are a greater strain)is important both in case of an existing pregnancy and in patientssuffering from HIV, leukaemia or under chemotherapy and markedly reducesthe strain on the patient, according to both subjective (compliance) andobjective criteria (strain on the metabolism). The chronic-recurrentvulvovaginitis in pregnancy can hardly be controlled by means of theconventional therapy regimen, since the systemic application of theorally applicable antimycotic agents (Fluconazol) is absolutelycounter-indicated in pregnancy.

Ad 2. Also in case of a chronic-recurrent course of the disease (inparticular in case of vulvovaginitis due to Candida infections), theunderlying pathogenic mechanism is interrupted, and a topical therapy isrendered possible instead of the long-lasting systemic therapy.

Ad 3. Compared to the hitherto usual therapy, the onset of the effect isgreatly accelerated, i.e. it occurs within minutes (vulvovaginitis) tohours (diaper thrush) as compared to days to weeks (or not at all) inconventional therapy.

Basically, the difference resides mainly in the duration of treatment,the therapy regimen proper will, however, first of all depend on thehalf-life of the pharmaceutical substances used. The difference may,e.g., be elucidated by way of the chronic-recurrent vulvovaginitis:

Use According to the Invention:

Initial therapy: 3-5× daily local application of the drug combination asan ointment, or by means of another locally applicable drug formulationfor three to five days;in case of recurrent affliction: 2-3× daily local application for oneday is sufficient (the recurrence-free intervals will always becomelonger with continued application).

In contrast, the present therapy regimen (source: Leitlinien derdeutschen Gesellschaft für Gynäkologie und Geburtshilfe):

Initial Therapy:

Fluconazol 150 mg orally, 1 or 2×/week for 4-6 weeks, subsequently

Fluconazol 150 mg orally, 1×/2 weeks for 2-3 months, subsequently

Fluconazol 150 mg orally, 1×/4 weeks for 4-6 months. Afterdiscontinuation, about 50% of the cases become recurrent, as before thetherapy.

Ad 4: Due to the potentiating effect, the total dose of the antimycoticcan be markedly reduced, in case of the uncomplicated vulvovaginalmycosis, one can assume a by at least 50% reduced total dose due to thereduced treatment period, in the oropharyngeal mycosis, the reductionmay be up to 66%.

Ad 5: With the often very painful acute clinical pictures, by thesimultaneous topical application of NSAID with the anti-mycotic agent,an immediate pain relief and detumescence with missing to minimal totalstrain on the organism is achieved with the NSAID (non-steroidalantiphlogistic agents) which, systemically applied, are not free fromside effects. As has been mentioned, the treatment according to theinvention is based on the finding that the expression of certainadhesion molecules at the tissue surface of the diseased organism ororgan, respectively, constitutes a basic prerequisite for thecolonization by yeasts. The combination of conventional antimycoticagents with epithelial cell or endothelial cell adhesion inhibitorstherefore directly interferes in the pathogenetic process. As has beenmentioned, epithelial cell or endothelial cell adhesion inhibitors aresubstances which exacerbate or prevent, respectively, the adhesion ofthe pathogens to epithelial and/or endothelial cells and the formationof a layer-shaped colonization of the afflicted epithelial orendothelial sector. From the nature of the responsible adhesionmolecules it results that their expression is prevented by activesubstances which interfere in the prostaglandin metabolism in a certainway. From the inventive combined application of antimycoticallyeffective substances and epithelial cell or endothelial cell adhesioninhibitors, there results a potentiation of the therapeutic effect. Theduration of treatment is greatly shortened, and a local treatmentbecomes possible also in those cases in which so far exclusively asystemic therapy had chances of success. Moreover, this type of acombination of pharmaceutical substances also is effective in caseswhich are completely therapy-resistant to drugs available at present.

Therefore, the subject matter of the invention is the use of theinventive combinations of pharmaceutical substances for the topicalapplication on mucosae and on greatly inflamed and/or macerated skinareas for the treatment of mycoses, preferably those caused by Candidaspp. (vulvovaginitis, oropharyngeal candidiasis (oral thrush), diaperdermatitis (diaper thrush), intertriginous eczema).

Application of the inventive combination preparation is exclusivelylocal (ointments, vaginal tablet, vaginal suppositories etc.) to skinand mucosae. The dosage of the anti-mycotic agent (preferablyClotrimazol) is effected at the concentration hitherto common, yet thedaily total dose can be halved, and the total treatment time is markedlyreduced (from 7 to 2-3 days). The epithelial cell or endothelial celladhesion inhibitor is administered at ⅕ to 1/100 of the daily maximumdose as an admixture to the ointment/vaginal tablet etc.

During the topical therapy with an inventive combination ofpharmaceutical substances, as mentioned above, the total amounts ofactive substance that are substantially lower than those hitherto usedin the antimycotic monotherapy suffice, and also the NSAID component isused at a significantly lower concentration than in all otherindications for which so far drugs of such substances have been used.Thus, also a treatment of breastfeeding babies and pregnant women ispossible.

Within a substantially shorter period of time (from minutes to a fewhours, as compared to from days to weeks when using the treatmentmethods common at present), a significant improvement of the subjectiveand objective disease symptoms will occur. This rapid onset of theeffect is solely caused by the processes at the surface of the mucosaand goes back to the effect of arachidonic acid derivatives triggered atthe afflicted organ surfaces and, therefore, can neither be detected inin-vitro tests nor by examinations carried out on the normal outer skin.

The nature of the antimycotic active substance to be employed in thepresent combination preparation in principle is not critical, as a rule,always an optimized combination pair will be used, wherein theoptimization primarily is based on the antimycotic spectrum of theantimycotic agent, on the galenics and on the physoco-chemicalinteractions of the anti-mycotic agent with the epithelial cell orendothelial cell adhesion inhibitor.

Preferred are, of course, those antimycotic agents which already have amarket authorization.

Therefore, the antimycotic agent in the combination preparationaccording to the invention preferably is an azole or a conazole,preferably Clotrimazole, Bifonazole, Croconazole, Miconazole, Econazole,Isoconazole, Itraconazole, Fenticonazole, Tioconazole, Sertaconazole,Sulconazole, Omoconazole, Oxiconazole, Fluconazole, Voriconazole orKetokonazole, in particular Clotrimazole and Miconazole, a squaleneepoxidase inhibitor, preferably Naftifin or Terbinafin, or anantibiotic, preferably Nystatin, Amphotericin B, Capsofungin orNatamycin, or Tolciclate, Tolnaftate, Ciclopirox, Haloprogin,Butenafine, Flucytosine.

Preferably, the combination drug according to the present invention isprepared as ointment, cream, lotion, gel, tincture, solution, vaginalsuppository, vaginal, buccal or sublingual tablet, syrup, suspension,powder, spray or aerosol.

According to a preferred embodiment of the present invention, thecombination drug may be provided on an inert carrier, in particular on avaginal ring, a diaphragm or a tampon.

The invention will be described in more detail by way of the followingexamples without, however, being restricted thereto.

Examinations and Results on Humans—Case Studies

So far, the therapy concept according to the invention has been examinedon voluntary subjects within the frame of the doctors' freedom ofprescription. Before the combination therapy, all the patients hadreceived an unsuccessful antimycotic monotherapy.

In the following, 6 examples will be described:

vulvovaginitis: 3 casesdiaper thrush: 2 casesoral thrush: 1 case

The treatment was by topical administration of suitable pharmaceuticalsubstance combinations on the skin, or on the mucosae of thegenitourinary tract and of the oropharynx, respectively.

The following combinations were used:

Vulvovaginitis:

Case 1:

A female 41-year old patient suffering from chronic-recurrent Candidavulvovaginitis for years (>10 recurrences/year), otherwise healthy,pregnant, prior treatment (before the pregnancy) both topical(Clotrimazole, Nystatin) and systemically (Fluconazole, several times,also long-term therapy and therapy of the partner). Massivedeterioration with complete therapy resistance to local treatment withClotrimazole during pregnancy, extreme subjective complaints for weeks.

Gyn. findings: vulva severely reddened, clearly swollen, bloodyexcoriations. Mucosa of the vagina and portio reddened. Massive vaginaldischarge, smear findings: PAP 2., microbial. swab, native finding:abundant leukocytes, fungus hyphae detectable in large masses, RG 3.

Therapy with an ointment combination of Clotrimazole/Diclofenac-Na

Single dose: 5 mg of Diclofenac-Na/20 mg of Clotrimazole Dosage regimen:initially (3 days) 3×/day ointment strips of 2.5 cm locally applied invulva and vagina, subsequently once per day, for 4 days.

The applied dose (total dose) of Diclofenac-Na is approximately 1/30 ofthe maximum daily dose at systemic application. The Clotrimazole dose isone half of the usual daily dose.

Findings after one week of therapy (three days after termination oftherapy):

Patient subjectively free from complaints, (since beginning of therapy).Gyn. findings: vulva: results negative; mucosae of vagina and portio:results negative. Low-grade vaginal discharge. Microbiol. swab, nativefindings: no fungus hyphae detectable, isolated spores. Normal vaginalflora (lactobacilli), RG 1.

Follow-up: 6 months

Over the next three months, approximately 2 recurrencies/month, one-daytreatment as above in each case resulted in immediate freedom fromcomplaints.Following months: complete freedom from complaints, no new recurrences.

Case 2:

43-year old female patient, suffering from chronic-recurrent Candidavulvovaginitis for years (>10 recurrences/year), otherwise healthy,prior treatment both topical (Clotrimazole, Miconazole) and systemically(Fluconazole, several times, also long-term therapy and therapy of thepartner). Acute exacerbation, low-grade improvement under therapy withClotrimazole (8 days), partly extreme subjective complaints for 2 weeks.

Gyn. findings: vulva severely reddened, clearly swollen. Mucosa of thevaginal and portio reddened. Massive vaginal discharge.

Initial therapy: Clotrimazole/Diclofenac (25/25 mg) supp. for 2 days,subsequently

Clotrimazole/Diclofenac as ointment as described in Case 1, for 3 days.

Findings after 1 week: patient subjectively free from complaints (since2^(nd) day after beginning of therapy). Gyn. findings: vulva resultsnegative; mucosae of vagina and portio: results negative. Low-gradevaginal discharge.

Case 3:

42-year old female patient, suffering from chronic-recurrent Candidavulvovaginitis for years (>10 recurrences/year), otherwise healthy,prior treatment both topical (Clotrimazole, Nystatin) and systemical(Fluconazole, several times, also long-term therapy and therapy of thepartner). Acute exacerbation, during this recurrence no pre-treatment,moderate subjective complaints for 2 days.

Gyn. findings: vulva reddened, swollen. Mucosa of the vaginal and portioreddened. Increased vaginal discharge.

Therapy: Clotrimazole/Diclofenac as ointment as described under Case 1on the first day 5×/day, from the 2^(nd) day onward 3×/day for a totalof three days.

Findings after 1 week: Patient subjectively free from complaints (since18 hours after the beginning of therapy). Gyn. findings: vulva resultsnegative; mucosae of vagina and portio: results negative. Low-gradevaginal discharge.

Case 4:

Diaper Thrush:

Female breastfeeding baby, 12 months old, therapy-resistant eczema formore than 5 weeks at the labia majora (pre-treatment by paediatricianand dermatologists with: zinc-oxide containing baby cream,Nystatin-containing cream, antibiotic (antibacterial) cream and powder,Clotrimazol-containing cream, corticosteroids. Under these therapies,continuous deterioration of the clinical picture.

Insp.: Pronounced reddening and swelling of the labia majora andperianally.

Therapy with an ointment combination of Clotrimazole/Diclofenac-Na

Single dose: ointment strips of approximately 5 cm in the evening(corresponding to approximately 10 mg of Diclofenac-Na/40 mgClotrimazole), complete healing over night. Subsequently, intestinalsanitation with Nystatin, oral suspension.

Follow-up (6 months): no further complaints

Case 5:

Diaper Thrush:

Female breastfeeding baby, 2 months old, repeated diaper thrush (redpapules).

Insp: Small papules on the skin of the labia majora and in the perianalregion.

Therapies tested: during various episodes, comparison of Clotrimazolecream with and without the addition of Diclofenac-Na.

Monotherapy: Clotrimazole cream (Canesten), single dose: approximately25 mg of ClotrimazoleCombination: Clotrimazole/Diclofenac-Na, single dose: approximately 5 mgof Diclofenac-Na/20 mg of Clotrimazole.In both cases, local application, dosage: 4-5×/day.

Result:

Monotherapy: duration of treatment until complete disappearance of thelesions: 3 days.Combination therapy: duration of treatment until complete disappearanceof lesions: 24 hours.

Case 6:

Oral Thrush:

Female breastfeeding baby, 2 months old, repeated oral thrush (whitelesions).

Insp.: typical white furs on the inner side of the upper and lower lips,diameter 4-5 mm.

Therapies tested: during various episodes comparison of Miconazole gelwith and without the addition of Mefenamic acid.

Monotherapy: Miconazole (Daktarin gel), single dose 30 mgCombination: Miconazole/mefenamic acid (gel).In both cases local application, dosage: 2-3×/daySingle dose: approximately 25 mg of mefenamic acid/30 mg of Miconazole

Result:

Monotherapy: duration of treatment until complete disappearance of thelesions: 5 daysCombination therapy: after 12 hours, significant reduction of thelesions, duration of treatment until complete disappearance of thelesions: 24 hours.

Summary of the Results of the Single Case Studies:

Vulvovaginitis: The cases described were particularly severe cases ofchronic-recurrent Candida vaginitis. Under monotherapy withClotrimazole, the course of curing was highly retarded in the secondcase (8 days of intensive therapy, then improvement, but no freedom fromcomplaints), in the first and third cases no improvement could beachieved by conventional local therapy with Clotrimazole. By using thepharmaceutical substance combinations, an immediate marked improvementof the subjective and objective symptoms occurred, and already on thesecond day of treatment, in all the cases the complaints completelydisappeared.

Diaper thrush: Under Clotrimazole monotherapy, marked deterioration ofthe clinical picture in the first case, healing after three days in thesecond (mild) case. Complete healing after the use of the pharmaceuticalsubstance combination within 12 and 24 hours, respectively.

Oral thrush: healing with Miconazole monotherapy after 4-5 days(criterion of diagnosis: no plaques visible any longer in the oralcavity). During a second episode, treatment with the pharmaceuticalsubstance combination, no plaques visible any longer after two hours.

These results show that in case of Candida mycoses which areparticularly difficult to treat, the combination preparation of theinvention brings about a surprisingly rapid and comprehensive healingwhich could not be achieved with the monotherapy (antimkycotic agentonly).

The success of the treatment was the clearer the more pronounced theinitial findings had been.

1.-6. (canceled)
 7. A method of treating a Candida mycosis furtherdefined as vulvovaginal candidiasis, oropharyngeal candidiasis (oralthrush), diaper dermatitis (diaper thrush), and/or intertriginous eczemacomprising: obtaining a combination drug comprising an antimycotic agentand an epithelial cell or endothelial cell adhesion inhibitor; andtopically treating a subject having a Candida mycosis further defined asvulvovaginal candidiasis, oropharyngeal candidiasis, diaper dermatitis,and/or intertriginous eczema with the combination drug.
 8. The method ofclaim 7, wherein the Candida mycosis is vulvovaginal candidiasis.
 9. Themethod of claim 7, wherein the Candida mycosis is oropharyngealcandidiasis.
 10. The method of claim 7, wherein the Candida mycosis isdiaper dermatitis.
 11. The method of claim 7, wherein the Candidamycosis is intertriginous eczema.
 12. The method of claim 7, wherein theantimycotic agent is an azole or conazole, a squalene epoxidaseinhibitor, or a polyene antimycotic agent.
 13. The method of claim 12,wherein the antimycotic agent is Clotrimazole, Bifonazole, Miconazole,Econazole, Isokonazole, Itraconazole, Fenticonazole, Tioconazole,Serticonazole, Omoconazole, Oxiconazole, Fluconazole, Naftifin,Terbinafin Nystatin, Amphotericin B, Capsofungin, Natamycin, Ciclopirox,Butenafine, or Flucytosin.
 14. The method of claim 7, wherein theepithelial cell or endothelial cell adhesion inhibitor is furtherdefined as: a non-steroidal antiphlogistic agent (NSAID) having asufficient COX1-inhibition, defined by the exclusion of compounds havinga COX1/COX2 ratio. (IC50, microM) of >20; a prostacyclin or aprostacyclin analogue; or an inhibitor of the expression of epithelialand endothelial adhesion molecules.
 15. The method of claim 14, whereinthe epithelial cell or endothelial cell adhesion inhibitor is Ilaprost,Cicaprost, Ticlopidine, or Clopidogrel.
 16. The method of claim 14,wherein the epithelial cell or endothelial cell adhesion inhibitor is anNSAID further defined as Indomethacin, mefenamic acid, Ketoprofen,acetyl salicylic acid, Ibuprofen, Lomoxicam, flufenamic acid,Diclofenac, Piroxicam, Bufexamac, Etofenamate, Felbinac, Tenoxicam, or apharmaceutically active salt or ester of any of these substances. 17.The method of claim 16, wherein the epithelial cell or endothelial celladhesion inhibitor is further defined as Diclofenac, Ibuprofen orLomoxicam.
 18. The method of claim 7, wherein the combination drug isfurther defined as an ointment, cream, lotion, gel, tincture, solution,vaginal suppository, vaginal, buccal or sublingual tablet, syrup,suspension, powder, spray, or aerosol.
 19. The method of claim 7,wherein the combination drug is comprised on an inert carrier.
 20. Themethod of claim 19, wherein the inert carrier is a vaginal ring,diaphragm, or tampon.